利用外周血单个核细胞生产成熟红细胞

体外制备红细胞的方法大多使用脐血、骨髓或外周动员的CD34＋细胞或胚胎干细胞作为起始材料.本研究利用外周血单个核细胞为原料在体外生产成熟红细胞.以血液滤涂白细胞后剩余的白膜层为原料,从中分离出单个核细胞,然后通过4步培养法制备成熟的红细胞.结果表明：经过不同细胞因子组合的诱导以及小鼠基质细胞系MS-5的支持培养,外周血单个核细胞的扩增倍数达到约1 000倍,其中约90％的细胞是与正常红细胞形态相似的无核成熟红细胞.集落形成能力分析显示,本研究培养体系可使单个核细胞中的祖细胞增殖并且只向红系方向分化.结构和功能分析结果表明,体外培养出的红细胞的平均细胞体积（MCV）为118±4fl,比正常红细胞（80-100 fl）稍大.平均血红蛋白含量（MCH）为36±1.2 pg,比正常的红细胞水平（27-31 pg）稍高.红细胞的最大变形指数（DImax）为0.46,接近正常红细胞水平.葡萄糖6磷酸脱氢酶及丙酮酸激酶的含量与年轻红细胞的特性一致.结论：利用外周血白膜层中含有的造血干祖细胞,通过体外培养能够生产出成熟红细胞,它是一种取材方便、成本低廉的红细胞生产原料,本研究培养体系适于利用单个核细胞生产成熟红细胞.

Production of Mature Red Blood Cell by Using Peripheral Blood Mononuclear Cells

Most protocols for in vitro producing red blood cells（RBC） use the CD34 ＋ cells or embryonic stem cells from cord blood,bone marrow or peripheral blood as the start materials.This study was purposed to produce the mature RBC in vitro by using peripheral blood mononuclear cells as start material.The peripheral blood mononuclear cells （PBMNC） were isolated from buffy coat after blood leukapheresis,the mature red blood ceils（RBC） were prepared by a 4-step culture protocol.The results showed that after culture by inducing with the different sets of cytokines and supporting by mouse MS-5 cell line,the expansion of PBMNC reached about 1000 folds at the end of the culture.About 90% of cultured RBC were enucleated mature cells which had the comparable morphological characteristics with normal RBC.Colony-forming assays showed that this culture system could stimulate the proliferation of progenitors in PBMNC and differentiate into erythroid cells.The structure and function analysis indicated that the mean cell volume of in vitro cultured RBC was 118 ±4 fl,which was slight larger than that of normal RBC （80-100 fl） ; the mean cell hemoglobin was 36 ± 1.2 pg,which was slight higher than that of normal RBC （27-31 pg） ; the maximal deformation index was 0.46,which approachs level of normal RBC; the glucose-6-phosphate dehydrogenase and pyrurvate kinase levels was consistant with young RBC.It is concluded that PBMNC are feasble,convenient and low-cost source for producing cultured RBC and this culture system is suitable to generate the RBC from PBMNC.